Dose-dependent induction of FOXO-GFP translocation by apigenin and luteolin, competition by insulin.

<p>Stably transfected human osteosarcoma cells with FOXO1-GFP (U2OS-FOXO1-GFP) treated with apigenin (A), luteolin (B) 1–100 µM for 2 h −/+ addition of insulin 100 nM after 30 minutes. Cells were fixed and stained with DAPI. Fluorescence microscopic detection of nuclei, segmentation of cells, quantification of GFP intensities measured in nuclear and cytoplasmic areas and calculation of the GFP-ratio nucleus/cytoplasm were performed for all FOXO-GFP expressing cells by BD Image Data Explorer. Nonlinear regression was performed with Graph Pad Prism. Results are presented as mean ± SEM of quadruplets with *(p<0.05) significant differences vs. control (ANOVA + Post Hoc Tests). (A) Dose-dependent accumulation of FOXO1 in nuclei induced by apigenin 1–100 µM shown as mean ratio of FOXO1 nucleus/cytoplasm + SEM (n = 4) *p<0.05 (Dunnett T3) and EC50 = 13 µM calculated by nonlinear regression from sigmoidal dose response. Reversion by insulin 100 nM induced FOXO1 translocation from nuclei into the cytoplasm. (B) Luteolin induced FOXO1 nuclear accumulation shown as mean ratio of FOXO1 nucleus/cytoplasm + SEM (n = 4) *p<0.05 (Bonferroni) with EC50 = 12 µM and competing insulin effect.